The long-time objective of this application is to understand the molecular biology of human postnatal stem cells from dental and periodontal tissues. Recently, human postnatal dental pulp stem cells (DPSCs) have been identified from dental pulp tissues. These cells are multipotent and can generate dentin/pulp-like complexes in vivo. Our exciting preliminary studies presented in this application also discovered that human periodontal ligament (PDL) contains unique stem cells (periodontal ligament stem cells; PDLSCs) which can generate a distinct cementum/PDL-like structure. These human postnatal dental and periodontal stem cells offer an attractive regenerative therapy for dental and periodontal defects caused by dental decay/pulpitis and periodontitis. However, before pushing them into clinical application, it is critical to develop optimal conditions to maintain their sternness during ex vivo expansion and to elucidate molecular mechanisms which control their differentiation and self-renewal. Like bone marrow mesenchymal stem cells (MSCs), these human postnatal dental stem cells appear to be from mesenchymal origin according to their surface markers. During in vitro culture, we also found that these stem cells, similar to MSCs, progressively lost their sternness. Based on our novel findings on the maintenance of MSCs' function by telomerase, in this application, we will examine whether over-expression of telomerases help to maintain human postnatal dental stem cell properties in vitro and whether Wnt growth factors stimulate telomerase activity and modulate their properties. Moreover, our preliminary studies suggest that Wnt signaling may negatively regulate the activation of NF-kappaB, a master transcription factor of inflammatory responses, and that NF-kappaB activated by inflammatory mediators such as TNF inhibit differentiation. Therefore, in this application, we will also explore whether Wnt signaling attenuates the inhibition of human dental stem cell differentiation by TNF. In the realm of therapeutic regeneration for dental and periodontal tissues, due to infection of oral pathogens, the repairing sites are frequently inflamed. Thus, our studies are highly clinically-relevant. Collectively, novel findings from our application will provide a molecular basis for maintaining and regulating human postnatal dental stem cell properties in vitro and in vivo, and have important implications in the regenerative dental medicine.